To find lead compounds for drug discovery programs, large numbers of compounds are often screened for their activity as enzyme inhibitors or receptor agonists/antagonists. Large libraries of compounds are needed for such screening. As a result of developments in this field, it is now possible to simultaneously produce combinatorial libraries containing hundreds of thousands of small molecules for screening. With the availability of such libraries, however, has come a need for large scale, rapid screening methods.
For example, the libraries may be contained on microbeads, each compound being present in a picomolar amount. Because the amount of compound is very small, it is advantageous to conduct the high throughput screening method in very small volumes, e.g., on the order of 1 .mu.l. Such assays can be performed in the 1536-well plate described in PCT Application Ser. No. PCT/US98/00494 entitled Multi-Well Plate, filed Jan. 8, 1998 and incorporated by reference herein.
Receptor binding assays used in high throughput screening typically involve three steps. First, a labeled ligand is incubated with a target receptor in the presence of a compound to be tested for inhibition of ligand/receptor binding. Second, the receptor and ligand (and compound) are separated using filtration and/or washing of an immobilized receptor. Finally, the amount of labelled ligand bound to the receptor is quantified. This conventional screening is a `separations-mode` assay, i.e., one in which the bound ligand is physically separated from the free ligand using either a filtration membrane or the selective adhesion of either bound or free component to a surface (e.g., the surface of a microtiter plate).
Separation, however, is typically time consuming and therefore slows high throughput screening. If fluid handling steps employed are not sufficiently precise, separation also can create variations in the signal generated in the assay and can disturb equilibrium binding conditions. Furthermore, separation has been difficult to automate and is potentially hazardous when radioactive materials are involved. These problems are particularly acute in assays conducted in microvolumes using small amounts of test compound.
Many screening assays require the transfer of cells or proteins onto membranes for evaluation of radiosotope levels, fluorescence or activity. For further details of various aspects of such assays, see U.S. patent application Ser. No. 08/868,280 filed Jun. 3, 1997, now U.S. Pat. No. 5,876,946, and entitled High-Throughput Assay which is incorporated by reference in its entirety herein. The small well size and volume used in the 1536-well plates preclude the use of conventional cell harvesting equipment.
For larger wells and lower throughput, Millipore provides a filtration system for use with 96-well format plates designated the MultiScreen.RTM. Assay System. In that system, individual membranes are sealed to 96 independent wells for use with a vacuum manifold. The membranes are relatively complex and expensive, and the vacuum manifold is relatively slow. Overall this system is not advantageously scaleable to the small volumes used with high throughput plates such as 1536-well plates.
Other conventional plate-to-plate transfer systems are inadequate at the small volumes used in 1536-well plates. Typical systems involve physical removal of the sample into a cannula or syringe, either with concomitant dispensing of a wash stream from a plate washer or in an aspirate/dispense mode. This modality is impractical using 1536-well plates due to the number of parallel operations that are needed to maintain an acceptably high throughput. In order to extend the prior art to 1536-well plates, a technique would need to be developed (at considerable expense) for fabricating parallel aspirate/dispense channels which might have disposable tips or washable cannulae to prevent contamination.
Thus, it would be highly desirable to have simpler, faster and more economical alternatives for transfers and separations of materials from a multiplicity of vessels, such as the wells of a microtiter plate. Further, to provide a rapid separation of cells or proteins from solutions within low volume high throughput plates, such as 1536-well plates, and to provide a mechanism for rapid plate-to-plate transfer of samples from one plate to another will be recognized as highly advantageous.